Development of Cryo-Electron Microscopy Sample Preparation for the Examination of Nanobubble

Cryo-electron microscopy (cryo-EM) has been widely used to characterize bio-macromolecules, such as DNA molecules, proteoliposomes, and protein complexes. The targets of interests were effectively remained the original structure and morphology in amorphous ice. However, in cryo-EM, target of interests in gas phase has not been studied yet. In this study, we utilized cryo-EM to examine the nanobubbles in frozen aqueous solution with a “sandwich” TEM grids assembly. With significantly increased surface area and lower buoyancy, nanobubble system greatly enhances the efficiency of oxygenation and the time of retention in medical application. The substrate of “sandwich” assembly provides sufficient nucleation site for the formation of bubbles. Additionally, elimination of water-air interface increases the possibility to capture bubbles in nano scale. This method demonstrates a promising way to evaluate bubble system using cryo-EM and provides an insight on the study of nanobubble in biomedical application.